AAVs (adeno-associated viruses) are single stranded DNA viruses belonging to the Parvovirus family. For their replication, i.e. for forming viral particles, AAVs require helper viruses, particularly adenoviruses or herpesviruses. In the absence of helper viruses, AAVs may incorporate into the host cell genome, particularly at a specific site of chromosome 19.
The genome of AAVs is linear and has a length of about 4680 nucleotides. It comprises two reading frames which code for a structural gene and a non-structural gene. The structural gene is referred to as cap gene. It is controlled by the P40 promoter and codes for three capsid proteins. The non-structural gene is referred to as rap gene and codes for the Rep proteins Rep 78, Rep 68, Rep 52 and Rep 40. The two former proteins are expressed under the control of the P5 promoter, while the expression of Rep 52 and Rep 40 is controlled by the P19 promoter. The functions of the Rep proteins are represented inter alia by the control of replication and transcription of the AAV genome.
It has now turned out that preparations of recombinant (r)AAV viral particles are frequently contaminated with helper viruses, e.g., adenoviruses or herpesviruses. This contamination considerably limits the use of rAAV viral particles for gene therapy. Efforts made to remove the helper viruses by CsC1 density gradient centrifugation or filtration methods have produced little success so far, in particular these methods comprise steps which manifest themselves negatively as regards costs and yield.
Therefore, it is the object of the present invention to provide a product by which rAAV viral particles can be provided without contamination with helper viruses.